We have developed a new approach to sensitively and quantitatively identify HIV-1-infected cells, and found that HIV-1 significantly infects some types of CD4(-) cells independently of gp120. Further studies have indicated that not only can CD4(-) cells be infected by HIV-1 through this mechanism, but also CD4(+) T-lymphocytes. These results propose two very important possibilities: 1) HIV-1 can exploit some types of CD4(-) cells to serve as viral reservoirs in patients; and 2) HIV-1 can reduce expression of Env in infected cells as a strategy to evade the host immune response and therefore reside in these cells as long-term low-level persistent infection in a latent state. In this application, we will quantitatively characterize gp120- independent HIV-1 infection, and elucidate the molecular mechanism by which this occurs. In Specific Aim 1, CD4(-) cell lines derived from various human tissues will be infected with three strains of HIV-1 Env(-) virus. The susceptibility of CD4(-) cells to HIV-1 infection, the rates of viral replication in infected cells, the life-span of the infected cells, and the expression of viral genes in these cell lines will be studied. The results from Aim 1 studies will be informative to assess the seriousness of overall HIV-1 infection of CD4(-) cells in patients. In Specific Aim 2, we will study the mechanism of gp120-independent infection. We will characterize the infection course of HIV-1 gp120- independent infection, including viral entry, reverse transcription, nuclear transportation, viral DNA integration, and the generation of viral RNA genomes using an HIV-susceptible CD4(-) cell line, LNCaP, infected by HIV-1 Env(-) virus. We will also identify genes that are involved in gp120-independent HIV-1 infection using subtractive hybridization. The results from these studies will be very informative for assessing the seriousness of CD4(-) cell hosted HIV-1 reservoirs in patients and for developing an approach to interrupt the infection of CD4(-) cells by HIV-1.